Clostridium difficile is one of the most problematic bacterial pathogens in developing countries with over 300,000 cases in the US annually. Cases of C. difficile are predicted to rise 40% over the next two years with billions of dollars of associated medical expenses. As antibiotic therapy often promotes colonization of C. difficile, more effective strategies to treat these infections are needed. Here, we propose to develop an innovative next generation Adenovirus (Ad) based vaccine targeting C. difficile combining technologies developed in the two PI's laboratories. Nonreplicating Ad based vaccines offer a unique, powerful vaccine delivery platform as genes encoding a desired antigen are directly expressed from the recombinant Ad vector. Our preliminary results developed a base Ad vaccine expressing a non-toxic portion of the C. difficile TA toxin that has shown promise as a vaccine candidate. However, to function as a therapeutic to rapidly reduce symptoms of individuals infected with C. difficile, generation of a rapid humoral response is required. One mechanism by which this can be achieved is the inclusion of an immunostimulatory adjuvant molecule into the vaccine. Here, we propose a novel approach to increase the efficacy of the previously constructed C. difficile Ad vaccine by inclusion of an adjuvant (specific details are excluded for protection of intellectual property). These experiments may lead to the development of a new vaccine for C. difficile. Additionally, our results will be proof-of-concept incorporation of this adjuvant can increase the efficacy of viral and DNA based vaccines to prevent infectious disease and other diseases, such as cancer.